DOI QR코드

DOI QR Code

In vitro analysis of antiviral immune response against avian influenza virus in chicken tracheal epithelial cells

  • Jubi Heo (Department of Animal Science and Technology, Chung-Ang University) ;
  • Thi Hao Vu (Department of Animal Science and Technology, Chung-Ang University) ;
  • CH Kim (Department of Animal Science and Technology, Chung-Ang University) ;
  • Anh Duc Truong (Department of Biochemistry and Immunology, National Institute of Veterinary Research) ;
  • Yeong Ho Hong (Department of Animal Science and Technology, Chung-Ang University)
  • 투고 : 2024.02.26
  • 심사 : 2024.05.20
  • 발행 : 2024.12.01

초록

Objective: Avian influenza virus (AIV) infections first affect the respiratory tract of chickens. The epithelial cells activate the host immune system, which leads to the induction of immune-related genes and the production of antiviral molecules against external environmental pathogens. In this study, we used chicken tracheal epithelial cells (TECs) in vitro model to investigate the immune response of the chicken respiratory tract against avian respiratory virus infections. Methods: Eighteen-day-old embryonic chicken eggs were used to culture the primary chicken TECs. Reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry (ICC) analysis of epithelial cell-specific gene makers were performed to confirm the characteristics, morphology, and growth pattern of primary cultured chicken TECs. Moreover, to investigate the cellular immune response to AIV infection or polyinosinic-polycytidylic acid (poly [I:C]) treatment, the TECs were infected with the H5N1 virus or poly (I:C). Then, immune responses were validated by RT-qPCR and western blotting. Results: The TECs exhibited polygonal morphology and formed colony-type cell clusters. The RT-qPCR results showed that H5N1 infection induced a significant expression of antiviral genes in TECs. We found that TECs treated with poly (I:C) and exposed to AIV infection-mediated activation of signaling pathways, leading to the production of antiviral molecules (e.g., pro-inflammatory cytokines and chemokines), were damaged due to the loss of junction proteins. We observed the activation of the nuclear factor kappa B and mitogen-activated protein kinase (MAPK) pathways, which are involved in inflammatory response by modulating the release of pro-inflammatory cytokines and chemokines in TECs treated with poly (I:C) and pathway inhibitors. Furthermore, our findings indicated that poly (I:C) treatment compromises the epithelial cell barrier by affecting junction proteins in the cell membrane. Conclusion: Our study highlights the utility of in vitro TEC models for unraveling the mechanisms of viral infection and understanding host immune responses in the chicken respiratory tract.

키워드

과제정보

This study was carried out with the support of the Cooperative Research Program for Agriculture Science and Technology Development (Project No. PJ015612), Rural Development Administration and Chung-Ang University Graduate Research Scholarship 2023, Republic of Korea.

참고문헌

  1. Alexander DJ. An overview of the epidemiology of avian influenza. Vaccine 2007;25:5637-44. https://doi.org/10.1016/j.vaccine.2006.10.051 
  2. Chan PKS. Outbreak of avian influenza A(H5N1) virus infection in Hong Kong in 1997. Clin Infect Dis 2002;34(Suppl 2):S58-64. https://doi.org/10.1086/338820 
  3. Swayne DE. Understanding the complex pathobiology of high pathogenicity avian influenza viruses in birds. Avian Dis 2007;51:242-9. https://doi.org/10.1637/7763-110706-regr.1 
  4. Ibricevic A, Pekosz A, Walter MJ, et al. Influenza virus receptor specificity and cell tropism in mouse and human airway epithelial cells. J Virol 2006;80:7469-80. https://doi.org/10.1128/JVI.02677-05 
  5. Iwasaki A, Foxman EF, Molony RD. Early local immune defences in the respiratory tract. Nat Rev Immunol 2017;17:7-20. https://doi.org/10.1038/nri.2016.117 
  6. Kato A, Schleimer RP. Beyond inflammation: airway epithelial cells are at the interface of innate and adaptive immunity. Curr Opin Immunol 2007;19:711-20. https://doi.org/10.1016/j.coi.2007.08.004 
  7. Wu NH, Yang W, Beineke A, et al. The differentiated airway epithelium infected by influenza viruses maintains the barrier function despite a dramatic loss of ciliated cells. Sci Rep 2016;6:39668. https://doi.org/10.1038/srep39668 
  8. Vu TH, Hong Y, Truong AD, et al. The highly pathogenic H5N1 avian influenza virus induces the mitogen-activated protein kinase signaling pathway in the trachea of two Ri chicken lines. Anim Biosci 2022;35:964-74. https://doi.org/10.5713/ab.21.0420 
  9. Kato A, Schleimer RP. Beyond inflammation: airway epithelial cells are at the interface of innate and adaptive immunity. Curr Opin Immunol 2007;19:711-20. https://doi.org/10.1016/j.coi.2007.08.004 
  10. Daidoji T, Kajikawa J, Arai Y, Watanabe Y, Hirose R, Nakaya T. Infection of human tracheal epithelial cells by H5 avian influenza virus is regulated by the acid stability of hemagglutinin and the pH of target cell endosomes. Viruses 2020;12:82. https://doi.org/10.3390/v12010082 
  11. Anderson JM. Molecular structure of tight junctions and their role in epithelial transport. Physiology 2001;16:126-30. https://doi.org/10.1152/physiologyonline.2001.16.3.126 
  12. Linfield DT, Raduka A, Aghapour M, Rezaee F. Airway tight junctions as targets of viral infections. Tissue Barriers 2021;9:1883965. https://doi.org/10.1080/21688370.2021.1883965 
  13. Short KR, Kasper J, van der Aa S, et al. Influenza virus damages the alveolar barrier by disrupting epithelial cell tight junctions. Eur Respir J 2016;47:954-66. https://doi.org/10.1183/13993003.01282-2015 
  14. Barjesteh N, O'Dowd K, Vahedi SM. Antiviral responses against chicken respiratory infections: Focus on avian influenza virus and infectious bronchitis virus. Cytokine 2020;127:154961. https://doi.org/10.1016/j.cyto.2019.154961 
  15. Lee J, Hong Y, Vu TH, et al. Influenza A pathway analysis of highly pathogenic avian influenza virus (H5N1) infection in genetically disparate Ri chicken lines. Vet Immunol Immunopathol 2022;246:110404. https://doi.org/10.1016/j.vetimm.2022.110404 
  16. Vu TH, Hong Y, Truong AD, et al. Cytokine-cytokine receptor interactions in the highly pathogenic avian influenza H5N1 virus-infected lungs of genetically disparate Ri chicken lines. Anim Biosci 2022;35:367-76. https://doi.org/10.5713/ab.21.0163 
  17. Esnault E, Bonsergent C, Larcher T, et al. A novel chicken lung epithelial cell line: characterization and response to low pathogenicity avian influenza virus. Virus Res 2011;159:32-42. https://doi.org/10.1016/j.virusres.2011.04.022 
  18. Barjesteh N, Taha-Abdelaziz K, Kulkarni RR, Sharif S. Innate antiviral responses are induced by TLR3 and TLR4 ligands in chicken tracheal epithelial cells: communication between epithelial cells and macrophages. Virology (Lond) 2019;534:132-42. https://doi.org/10.1016/j.virol.2019.06.003 
  19. Fleming SB. Viral inhibition of the IFN-induced JAK/STAT signalling pathway: development of live attenuated vaccines by mutation of viral-encoded IFN-antagonists. Vaccines (Basel) 2016;4:23. https://doi.org/10.3390/vaccines4030023 
  20. Vu TH, Heo J, Hong Y, et al. HPAI-resistant Ri chickens exhibit elevated antiviral immune-related gene expression. J Vet Sci 2023;24:e13. https://doi.org/10.4142/jvs.22229 
  21. Haller O, Staeheli P, Schwemmle M, Kochs G. Mx GTPases: dynamin-like antiviral machines of innate immunity. Trends Microbiol 2015;23:154-63. https://doi.org/10.1016/j.tim.2014.12.003 
  22. Melchjorsen J, Kristiansen H, Christiansen R, et al. Differential regulation of the OASL and OAS1 genes in response to viral infections. J Interferon Cytokine Res 2009;29:199-208. https://doi.org/10.1089/jir.2008.0050 
  23. Kaminska B. MAPK signalling pathways as molecular targets for anti-inflammatory therapy-from molecular mechanisms to therapeutic benefits. Biochim Biophys Acta Proteins Proteom 2005;1754:253-62. https://doi.org/10.1016/j.bbapap.2005.08.017 
  24. Xing Z, Cardona CJ, Anunciacion J, Adams S, Dao N. Roles of the ERK MAPK in the regulation of proinflammatory and apoptotic responses in chicken macrophages infected with H9N2 avian influenza virus. J Gen Virol 2010;91:343-51. https://doi.org/10.1099/vir.0.015578-0 
  25. Pan H, Zhang Y, Luo Z, et al. Autophagy mediates avian influenza H5N1 pseudotyped particle-induced lung inflammation through NF-kappaB and p38 MAPK signaling pathways. Am J Physiol Lung Cell Mol Physiol 2014;306:L183-95. https://doi.org/10.1152/ajplung.00147.2013 
  26. Craig R, Larkin A, Mingo AM, et al. p38 MAPK and NF-κB collaborate to induce interleukin-6 gene expression and release: evidence for a cytoprotective autocrine signaling pathway in a cardiac myocyte model system. J Biol Chem 2000;275:23814-24. https://doi.org/10.1074/jbc.M909695199 
  27. Park JW, Ndimukaga M, So J, et al. Molecular analysis of chicken interferon-alpha inducible protein 6 gene and transcriptional regulation. J Anim Sci Technol 2023;65:183-96. https://doi.org/10.5187/jast.2022.e101 
  28. Gao N, Rezaee F. Airway epithelial cell junctions as targets for pathogens and antimicrobial therapy Pharmaceutics 2022;14:2619. https://doi.org/10.3390/pharmaceutics14122619 
  29. Ruan T, Sun Y, Zhang J, et al. H5N1 infection impairs the alveolar epithelial barrier through intercellular junction proteins via Itch-mediated proteasomal degradation. Commun Biol 2022;5:186. https://doi.org/10.1038/s42003-022-03131-3 
  30. Wei J, Jiang H, Gao H, Wang G. Activation of toll like receptor-3 induces corneal epithelial barrier dysfunction. Biochem Biophys Res Commun 2015;461:555-9. https://doi.org/10.1016/j.bbrc.2015.04.080 
  31. Tata PR, Mou H, Pardo-Saganta A, et al. Dedifferentiation of committed epithelial cells into stem cells in vivo. Nature 2013;503:218-23. https://doi.org/10.1038/nature12777 
  32. Rezaee F, Meednu N, Emo JA, et al. Polyinosinic: polycytidylic acid induces protein kinase D-dependent disassembly of apical junctions and barrier dysfunction in airway epithelial cells. J Allergy Clin Immunol 2011;128:1216-24. https://doi.org/10.1016/j.jaci.2011.08.035